The ability of cancer to develop resistance to therapy is a major health problem which was identified recently by the NCI Cancer Moonshoot initiative as a priority research topic. A critical barrier to progress in dealing with the problem of cancer therapy is the lack of drugs that reverse therapeutic resistance and block the supportive actions of the tumor microenvironment. Our previous research began addressing this need by identifying the autotaxin (ATX)?lysophosphatidic acid (LPA) axis as a fundamental system that regulates therapeutic resistance in various cancer stem-like cells (CSC) and mediates metastasis by reprogramming cells in the tumor microen- vironment (TME); and we developed drug candidate leads that inhibit ATX, which we will advance toward human trials. The next phase of our studies will expedite clinical availability and FDA approval of our lead ATX inhibitors. Our goal is to address cancer resistance to therapy by determining the mechanism of ATX upregulation in cells of the tumor microenvironment (TME) and to evaluate the therapeutic utility of four high-potency small-molecule inhibitors of ATX that we have developed during the previous funding period. Our central hypothesis is that transcriptional upregulation of ATX is responsible for the development of thera- peutic resistance by CSC and for reprogramming cells of the TME to provide a niche that is more conducive to invasion and metastasis. Our preliminary data suggest that our small-molecule ATX inhibitor leads will restore therapeutic sensitivity of CSC and also block ATX in the TME, which will enhance tumor cell killing and prevent the spread of cancer. Our objectives are: 1) Determine the cellular sources of ATX in the TME to pave the way to cell-targeted therapy against metastasis. 2) Develop an effective drug that simultaneously suppresses ATX in cancer stem cells proper and also in cells of the TME. This dual targeting will hit neoplastic cells at two of the most problematic areas of current cancer therapy: re- sistance and metastasis. Our expected outcomes will include: 1) validation that ATX is a new target in cancer treatment; 2) identification of the cellular targets that are sources of ATX in the TME; 3) identification of the signaling pathway of ATX upregulation in cancer-associated fibroblasts; and 4) evaluation of four new ATX inhibitor leads as a first-in-class drug candidate family. Our impact will include the preclinical validation of the most efficacious ATX inhibitor compound among our leads for submission in the NCI I-SPY Phase 1 human trial program as adjuvants to chemo- and radiation ther- apy. Our mechanism of action studies will also identify additional new drug targets that can block ATX expres- sion. We vigorously will seek industry partnerships/licensing to expedite clinical trials and regulatory approval.